A Small Molecule RAS-Mimetic Disrupts RAS Association with Effector Proteins to Block Signaling.

Oncogenic activation of RAS genes via point mutations occurs in 20%-30% of human cancers. The development of effective RAS inhibitors has been challenging, necessitating new approaches to inhibit this oncogenic protein. Functional studies have shown that the switch region of RAS interacts with a large number of effector proteins containing a common RAS-binding domain (RBD). Because RBD-mediated interactions are essential for RAS signaling, blocking RBD association with small molecules constitutes an attractive therapeutic approach. Here, we present evidence that rigosertib, a styryl-benzyl sulfone, acts as a RAS-mimetic and interacts with the RBDs of RAF kinases, resulting in their inability to bind to RAS, disruption of RAF activation, and inhibition of the RAS-RAF-MEK pathway. We also find that ribosertib binds to the RBDs of Ral-GDS and PI3Ks. These results suggest that targeting of RBDs across multiple signaling pathways by rigosertib may represent an effective strategy for inactivation of RAS signaling.

A Contaminant Impurity, Not Rigosertib, Is a Tubulin Binding Agent.

Rigosertib is a styryl benzyl sulfone that inhibits growth of tumor cells and acts as a RAS mimetic by binding to Ras binding domains of RAS effectors. A recent study attributed rigosertib's mechanism of action to microtubule binding. In that study, rigosertib was obtained from a commercial vendor. We compared the purity of clinical-grade and commercially sourced rigosertib and found that commercially sourced rigosertib contains approximately 5% ON01500, a potent inhibitor of tubulin polymerization. Clinical-grade rigosertib, which is free of this impurity, does not exhibit tubulin-binding activity. Cell lines expressing mutant ß-tubulin have also been reported to be resistant to rigosertib. However, our study showed that these cells failed to proliferate in the presence of rigosertib at concentrations that are lethal to wild-type cells. Rigosertib induced a senescence-like phenotype in the small percentage of surviving cells, which could be incorrectly scored as resistant using short-term cultures.

Duration of Adjunctive Antidepressant Maintenance in Bipolar I Depression.

BACKGROUND: Antidepressants are used to treat acute depression in patients with bipolar I disorder, but their effect as maintenance treatment after the remission of depression has not been well studied. METHODS: We conducted a multisite, double-blind, randomized, placebo-controlled trial of maintenance of treatment with adjunctive escitalopram or bupropion XL as compared with discontinuation of antidepressant therapy in patients with bipolar I disorder who had recently had remission of a depressive episode. Patients were randomly assigned in a 1:1 ratio to continue treatment with antidepressants for 52 weeks after remission or to switch to placebo at 8 weeks. The primary outcome, assessed in a time-to-event analysis, was any mood episode, as defined by scores on scales measuring symptoms of hypomania or mania, depression, suicidality, and mood-episode severity; additional treatment or hospitalization for mood symptoms; or attempted or completed suicide. Key secondary outcomes included the time to an episode of mania or hypomania or depression. RESULTS: Of 209 patients with bipolar I disorder who participated in an open-label treatment phase, 150 who had remission of depression were enrolled in the double-blind phase in addition to 27 patients who were enrolled directly. A total of 90 patients were assigned to continue treatment with the prescribed antidepressant for 52 weeks (52-week group) and 87 were assigned to switch to placebo at 8 weeks (8-week group). The trial was stopped before full recruitment was reached owing to slow recruitment and funding limitations. At 52 weeks, 28 of the patients in the 52-week group (31%) and 40 in the 8-week group (46%) had a primary-outcome event. The hazard ratio for time to any mood episode in the 52-week group relative to the 8-week group was 0.68 (95% confidence interval [CI], 0.43 to 1.10; P = 0.12 by log-rank test). A total of 11 patients in the 52-week group (12%) as compared with 5 patients in the 8-week group (6%) had mania or hypomania (hazard ratio, 2.28; 95% CI, 0.86 to 6.08), and 15 patients (17%) as compared with 35 patients (40%) had recurrence of depression (hazard ratio, 0.43; 95% CI, 0.25 to 0.75). The incidence of adverse events was similar in the two groups. CONCLUSIONS: In a trial involving patients with bipolar I disorder and a recently remitted depressive episode, adjunctive treatment with escitalopram or bupropion XL that continued for 52 weeks did not show a significant benefit as compared with treatment for 8 weeks in preventing relapse of any mood episode. The trial was stopped early owing to slow recruitment and funding limitations. (Funded by the Canadian Institutes of Health Research; ClinicalTrials.gov number, NCT00958633.).

Identification of candidate IgG biomarkers for Alzheimer's disease via combinatorial library screening.

The adaptive immune system is thought to be a rich source of protein biomarkers, but diagnostically useful antibodies remain unknown for a large number of diseases. This is, in part, because the antigens that trigger an immune response in many diseases remain unknown. We present here a general and unbiased approach to the identification of diagnostically useful antibodies that avoids the requirement for antigen identification. This method involves the comparative screening of combinatorial libraries of unnatural, synthetic molecules against serum samples obtained from cases and controls. Molecules that retain far more IgG antibodies from the case samples than the controls are identified and subsequently tested as capture agents for diagnostically useful antibodies. The utility of this method is demonstrated using a mouse model for multiple sclerosis and via the identification of two candidate IgG biomarkers for Alzheimer's disease.

Network Pharmacology Approach to Identify the Calotropis Phytoconstituents' Potential Epileptic Targets and Evaluation of Molecular Docking, MD Simulation, and MM-PBSA Performance.

Epilepsy originates from unusual electrical rhythm within brain cells, causes seizures. Calotropis species have been utilized to treat a wide spectrum of ailments since antiquity. Despite chemical and biological investigations, there have been minimal studies on their anticonvulsant activity, and the molecular targets of this plant constituents are unexplored. This study aimed to investigate the plausible epileptic targets of Calotropis phytoconstituents through network pharmacology, and to evaluate their binding strength and stability with the identified targets. In detail, 125 phytoconstituents of the Calotropis plant (C. procera and C. gigantea) were assessed for their drug-likeness (DL), blood-brain-barrier (BBB) permeability and oral bioavailability (OB). Network analysis revealed that targets PTGS2 and PPAR-γ were ranked first and fourth, respectively, among the top ten hub genes significantly linked with antiepileptic drug targets. Additionally, docking, molecular dynamic (MD) simulation, and Molecular Mechanics-Poisson-Boltzmann Surface Area (MM-PBSA) were employed to validate the compound-gene interactions. Docking studies suggested ergost-5-en-3-ol, stigmasterol and ß-sitosterol exhibit stronger binding affinity and favorable interactions than co-crystallized ligands with both the targets. Furthermore, both MD simulations and MM-PBSA calculations substantiated the docking results. Combined data revealed that Calotropis phytoconstituents ergost-5-en-3-ol, stigmasterol, and ß-sitosterol might be the best inhibitors of both PTGS2 and PPAR-γ.

Unraveling different begomoviruses, DNA satellites and cryptic species of Bemisia tabaci and their endosymbionts in vegetable ecosystem.

Bemisia tabaci species complex contains more than 46 cryptic species. It has emerged as an important pest causing significant yield loss in many cultivated crops. This pest is also a vector for more than 100 species of begomoviruses, that are a major threat for the cultivation of many crops in different regions of the world. The relation between cryptic species of the B. tabaci species complex and associated begomoviruses that infect different crops remains unclear. In the present study, four cryptic species (Asia I, China 3, Asia II 5 and Asia II-1) of B. tabaci and four associated endosymbionts (Arsenophonus, Cardinium, Rickettsia and Wolbachia) were identified in different vegetable crops. The vector-based PCR detection revealed five different begomoviruses such as okra enation leaf curl virus (OELCuV), tomato leaf curl Palampur virus (ToLCPalV), squash leaf curl China virus (SLCCNV), chilli leaf curl virus (ChiLCuV), and tomato leaf curl New Delhi virus (ToLCNDV). Of these begomoviruses, the maximum infection rate was observed (9.1%) for OELCuV, followed by 7.3% for ToLCNDV. The infection rate of the other three viruses (SLCCNV, ChiLCuV, ToLCPalV) ranged from 0.9 to 2.7% in cryptic species of B. tabaci. Further, each cryptic species was infected with multiple virus species and the virus infection rate of Asia I, Asia II-5, China 3 and Asia II-1 was 21.2%, 15.1%, 15.1% and 0.6% respectively. Similarly, in case of betasatellites the highest infection rate was 12% for ToLCBDB, followed by 6% for OLCuB and PaLCB. With regard to alphasatellites, the highest infection rate was 18.2% for AEV and 3% for CLCuMuA. This study demonstrates the distribution of cryptic species of whitefly and their endosymbionts, and associated begomoviruses and DNA satellites in vegetable ecosystem. We believe that the information generated here is useful for evolving an effective pest management strategies for vegetable production.

Diversity and phylogeography of begomoviruses and DNA satellites associated with the leaf curl and mosaic disease complex of eggplant.

Eggplant is one of the important vegetable crops grown across the world, and its production is threatened by both biotic and abiotic stresses. Diseases caused by viruses are becoming major limiting factors for its successful cultivation. A survey for begomovirus-like symptoms in 72 eggplant fields located in six different Indian states revealed a prevalence of disease ranging from 5.2 to 40.2%, and the symptoms recorded were mosaic, mottling, petiole bending, yellowing, and upward curling, vein thickening, and enation of the leaves, and stunting of plants. The causal agent associated with these plants was transmitted from infected leaf samples to healthy eggplant seedlings via grafting and whiteflies (Bemisia tabaci). The presence of begomovirus was confirmed in 72 infected eggplant samples collected from the surveyed fields exhibiting leaf curl and mosaic disease by PCR using begomovirus specifc primers (DNA-A componet), which resulted in an expected amplicon of 1.2 kb. The partial genome sequence obtained from amplified 1.2 kb from all samples indicated that they are closely related begomovirus species, tomato leaf Karnataka virus (ToLCKV, two samples), tomato leaf curl Palampur virus (ToLCPalV, fifty eggplant samples), and chilli leaf curl virus (ChLCuV, twenty samples). Based on the partial genome sequence analysis, fourteen representative samples were selected for full viral genome amplification by the rolling circle DNA amplification (RCA) technique. Analyses of fourteen eggplant isolates genome sequences using the Sequence Demarcation Tool (SDT) indicated that one isolate had the maximum nucleotide (nt) identity with ToLCKV and eight isolates with ToLCPalV. Whereas, four isolates four isolates (BLC1-CH, BLC2-CH, BLC3-CH, BLC4-CH) are showing nucleotide identity of less than 91% with chilli infecting viruses begomoviruses with chilli infecting begomoviruses and as per the guidelines given by the ICTV study group for the classification of begomoviruses these isolates are considered as one novel begomovirus species, for which name, Eggplant leaf curl Chhattisgarh virus (EgLCuChV) is proposed. For DNA-B component, seven eggplant isolates had the highest nt identity with ToLCPalV infecting other crops. Further, DNA satellites sequence analysis indicated that four betasatellites identified shared maximum nucleotide identity with the tomato leaf curl betasatellite and five alphasatellites shared maximum nucleotide identity with the ageratum enation alphasatellite. Recombination and GC plot analyses indicated that the bulk of begomovirus genome and associated satellites presumably originated from of previously known mono and bipartite begomoviruses and DNA satellites. To the best of our knowledge, this is India's first report of ToLCKV and a noval virus, eggplant leaf curl Chhattisgarh virus associated with eggplant leaf curl disease.

Direct method to grasp molecular topology of mesogens through 13C-1H dipolar couplings.

A prime factor in determining liquid crystalline phase formation is the overall molecular shape since molecules undergo rotational motion about the long axis. Molecular topology deals with the connectivity of atomic centers in a given molecular architecture, ultimately giving rise to the gross molecular shape. 13C NMR has emerged as the most important technique in establishing the molecular topology of mesogens in the liquid crystalline phase. In this work, we demonstrate the utility of 13C-1H dipolar couplings determined from 2D separated local field NMR for finding the topology of three different mesogens in the liquid crystalline phase. The core unit of the investigated mesogens fundamentally differs, which may be categorized as rod-like, laterally substituted, and bent-core shapes. 1D and 2D 13C NMR measurements in the liquid crystalline phase revealed fascinating information. The 13C-1H dipolar couplings extracted from 2D NMR are found to be sensitive to topologically variant core units. This permitted us to establish the molecular topology just by looking at the 13C-1H dipolar couplings of the protonated carbons of the constituent phenyl rings. By considering the dipolar couplings of rod-like mesogens as a reference, the large variation in the magnitude of 13C-1H dipolar couplings of the laterally substituted and bent-core mesogens is attributed to changes in the topology of their core units. The order parameters estimated from 13C-1H dipolar couplings enabled visualization of the ordering array of phenyl rings of the mesogens. Interestingly large 13C-1H dipolar couplings are observed for mesogens in which (a) laterally located phenyl ring and (b) central phenyl ring of bent-core mesogens exhibited different trends as revealed by the orientational order parameters.

Acceptable intakes (AIs) for 11 small molecule N-nitrosamines (NAs).

Low levels of N-nitrosamines (NAs) were detected in pharmaceuticals and, as a result, health authorities (HAs) have published acceptable intakes (AIs) in pharmaceuticals to limit potential carcinogenic risk. The rationales behind the AIs have not been provided to understand the process for selecting a TD50 or read-across analog. In this manuscript we evaluated the toxicity data for eleven common NAs in a comprehensive and transparent process consistent with ICH M7. This evaluation included substances which had datasets that were robust, limited but sufficient, and substances with insufficient experimental animal carcinogenicity data. In the case of robust or limited but sufficient carcinogenicity information, AIs were calculated based on published or derived TD50s from the most sensitive organ site. In the case of insufficient carcinogenicity information, available carcinogenicity data and structure activity relationships (SARs) were applied to categorical-based AIs of 1500 ng/day, 150 ng/day or 18 ng/day; however additional data (such as biological or additional computational modelling) could inform an alternative AI. This approach advances the methodology used to derive AIs for NAs.

Rate of Recovery and Symptomatic Efficacy of a Polyherbal AYUSH Formulation in the Treatment of SARS-CoV-2 Disease: A Single-arm Trial.

Background: The COVID-19 pandemic, caused by the human coronavirus, SARS-CoV-2, has led to the death of millions across the globe. The SARS-CoV-2 virus is highly infectious, and mutates rapidly. This creates additional challenges for the development of robust therapeutic solutions. Along with modern healthcare, there is a need to explore natural, plant-based antiviral compounds that can be used in the treatment of COVID-19. Objective: The present feasibility study investigates the efficacy of a 13-ingredient Ayurvedic polyherbal formulation, NOQ19, in the management of COVID-19. Methodology: A single-arm, open-label study design was adopted for this feasibility study. 161 RT-PCR-positive COVID-19 patients were enrolled. The enrolled participants were provided with the Ayurvedic intervention - two tablets of NOQ19 thrice daily along with the standard of care treatment. Follow-up COVID-19 RT-PCR tests were conducted on days 5, 10, and 14 or until the patient tested negative. The time taken to turn RT-PCR negative or become asymptomatic was noted. Setting: The study was conducted at Sri Sri Institute for Advanced Research from April 2021 to June 2021. Participants: A total of 161 COVID-19 patients isolating at home were assessed. Intervention: The NOQ19 preparation is a combination of 13 Ayurvedic herbs. Outcomes Measured: RT-PCR tests, the turnaround time to becoming asymptomatic, and regular symptoms assessment. Results: The analysis demonstrated that 74% of the patients tested negative on the RT-PCR within five days of taking NOQ19. Additionally, 98% of the subjects tested negative on the RT-PCR on day 10 after taking NOQ19 and standard of care treatment (as necessary). None of the participants reported any adverse events or side-effects due to NOQ19 medication. Conclusion: The NOQ19 Ayurvedic polyherbal formulation can be an effective and safe option for the symptomatic management of COVID-19.

Removal of cadmium and arsenic from water through biomineralization.

Due to anthropogenic activities, heavy metals such as cadmium (Cd) and arsenic (As) are one of the most toxic xenobiotics contaminating water, thus affecting human health and the environment. The objective of the present investigation was to study the effect of ureolytic bacteria Bacillus paramycoides-MSR1 for the bioremediation of Cd and As from contaminated water. The B. paramycoides showed high resistance to heavy metals, Cd and As, with minimum inhibitory concentration (MIC) of 12.84 µM and 48.54 µM, respectively. The urease activity and calcium carbonate (CaCO3) precipitation were evaluated in artificial wastewater with different concentrations of Cd (0, 10, 20, 30, 40, 50, and 60 µM) and As (0, 20, 40, 60, 80, and 100 µM). The maximum urease activity in Cd-contaminated artificial wastewater was observed after 96 hours, which showed a 76.1% decline in urease activity as the metal concentration increased from 0 to 60 µM. Similarly, 14.1% decline in urease activity was observed as the concentration of As was increased from 0 to 100 µM. The calcium carbonate precipitation at the minimum inhibitory concentration of Cd and As-contaminated artificial wastewater was 189 and 183 mg/100 ml, respectively. The percentage removal of metal from artificially contaminated wastewater with varied concentrations was analyzed using atomic absorption spectroscopy (AAS). After 168 hours of incubation, 93.13% removal of Cd and 94.25% removal of As were observed. Microstructural analysis proved the presence of calcium carbonate in the form of calcite, confirming removal of cadmium and arsenic by microbially induced calcium carbonate precipitation (MICCP) to be promising technique for water decontamination.

Role of placental, fetal and maternal cardiovascular markers in predicting adverse outcome in women with suspected or confirmed pre-eclampsia.

OBJECTIVE: To assess the performance of placental, fetal and maternal cardiovascular markers in the prediction of adverse perinatal and maternal outcomes in women with suspected or confirmed pre-eclampsia. METHODS: This was a prospective prognostic accuracy study of women with suspected or confirmed pre-eclampsia who underwent a series of investigations to measure maternal hemodynamic indices, mean arterial pressure, augmentation index, ophthalmic artery peak systolic velocity (PSV) ratio, uterine artery pulsatility index (UtA-PI), fetal biometric and Doppler parameters, soluble fms-like tyrosine kinase-1 (sFlt-1) and placental growth factor (PlGF). The performance of these markers, individually or in combination, in predicting adverse perinatal and maternal outcomes was then assessed using receiver-operating-characteristics (ROC)-curve analysis. Adverse maternal outcome was defined as one or more of severe hypertension, admission to the intensive care unit, eclampsia, placental abruption, HELLP syndrome, disseminated intravascular coagulation, platelets < 100 × 109 /L, creatinine > 90 µmol/L and alanine aminotransferase > 100 U/L. Adverse perinatal outcome was defined as one or more of preterm birth at or before 34 + 0 weeks, neonatal intensive care unit admission for > 48 h, respiratory distress syndrome, intraventricular hemorrhage, hypoxic ischemic encephalopathy, necrotizing enterocolitis, retinopathy of prematurity and confirmed fetal infection. RESULTS: We recruited 126 women with suspected (n = 31) or confirmed (n = 95) pre-eclampsia at a median gestational age of 33.9 weeks (interquartile range, 30.9-36.3 weeks). Pregnancies with adverse perinatal outcome compared to those without had a higher median UtA-PI (1.3 vs 0.8; P < 0.001), ophthalmic artery PSV ratio (0.8 vs 0.7; P = 0.01) and umbilical artery PI percentile (82.0 vs 68.5; P < 0.01) and lower median estimated fetal weight percentile (4.0 vs 43.0; P < 0.001), abdominal circumference percentile (4.0 vs 63.0; P < 0.001), middle cerebral artery PI percentile (28.0 vs 58.5; P < 0.001) and cerebroplacental ratio percentile (18.0 vs 46.5; P < 0.001). Pregnancies with adverse perinatal outcome also had a higher median sFlt-1 (8208.0 pg/mL vs 4508.0 pg/mL; P < 0.001), lower PlGF (27.2 pg/mL vs 76.3 pg/mL; P < 0.001) and a higher sFlt-1/PlGF ratio (445.4 vs 74.4; P < 0.001). The best performing individual marker for predicting adverse perinatal outcome was the sFlt-1/PlGF ratio (area under the ROC curve (AUC), 0.87 (95% CI, 0.81-0.93)), followed by estimated fetal weight (AUC, 0.81 (95% CI, 0.73-0.89)). Women who experienced adverse maternal outcome had a higher median sFlt-1 level (7471.0 pg/mL vs 5131.0 pg/mL; P < 0.001) and sFlt-1/PlGF ratio (204.3 vs 93.3; P < 0.001) and a lower PlGF level (37.0 pg/mL vs 66.1 pg/mL; P = 0.01) and estimated fetal weight percentile (16.5 vs 37.0; P = 0.04). All markers performed poorly in predicting adverse maternal outcome, with sFlt-1 (AUC, 0.69 (95% CI, 0.60-0.79)) and sFlt-1/PlGF ratio (AUC, 0.69 (95% CI, 0.59-0.78)) demonstrating the best individual performance. The addition of cardiovascular, fetal or other placental indices to the sFlt-1/PlGF ratio did not improve the prediction of adverse maternal or perinatal outcomes. CONCLUSIONS: The sFlt-1/PlGF ratio performs well in predicting adverse perinatal outcomes but is a poor predictor of adverse maternal outcomes in women with suspected or diagnosed pre-eclampsia. The addition of cardiovascular or fetal indices to the model is unlikely to improve the prognostic performance of the sFlt-1/PlGF ratio. © 2022 International Society of Ultrasound in Obstetrics and Gynecology.

Knowledge extraction of sonophotocatalytic treatment for acid blue 113 dye removal by artificial neural networks.

Removing decolorizing acid blue 113 (AB113) dye from textile wastewater is challenging due to its high stability and resistance to removal. In this study, we used an artificial neural network (ANN) model to estimate the effect of five different variables on AB113 dye removal in the sonophotocatalytic process. The five variables considered were reaction time (5-25 min), pH (3-11), ZnO dosage (0.2-1.0 g/L), ultrasonic power (100-300 W/L), and persulphate dosage (0.2-3 mmol/L). The most effective model had a 5-7-1 architecture, with an average deviation of 0.44 and R2 of 0.99. A sensitivity analysis was used to analyze the impact of different process variables on removal efficiency and to identify the most effective variable settings for maximum dye removal. Then, an imaginary sonophotocatalytic system was created to measure the quantitative impact of other process parameters on AB113 dye removal. The optimum process parameters for maximum AB 113 removal were identified as 6.2 pH, 25 min reaction time, 300 W/L ultrasonic power, 1.0 g/L ZnO dosage, and 2.54 mmol/L persulfate dosage. The model created was able to identify trends in dye removal and can contribute to future experiments.

Living Donor Liver Transplantation for Budd-Chiari Syndrome: A Propensity Score-Matched Analysis.

INTRODUCTION: There are unique technical and management challenges associated with living donor liver transplantation (LDLT) for Budd-Chiari Syndrome (BCS). The outcomes of LDLT for BCS in comparison to other indications remain unclear and warrant elucidation. METHODS: Data of 24 BCS patients who underwent LDLT between January 2012 and June 2019 were analyzed. There were 20 adults and 4 children. The early and long-term outcomes of adult LDLT BCS patients were compared to a control group of LDLT patients for other indications and matched using propensity scoring methodology. RESULTS: Primary BCS was observed in 18 (90%) patients. Caval replacement was performed in 7 (35%) patients. Early and late hepatic venous outflow tract (HVOT) complications were seen in 1 (5%) and 3 (15%) patients. Preoperative acute kidney injury was identified as a risk factor for mortality in the BCS cohort (p = 0.013). On comparison, BCS recipients were younger with fewer comorbidities, more large volume ascites and higher rates of PVT. They also had longer cold ischemia time, increased blood loss and transfusion requirements, increased hospital stay, and higher late outflow complications. The 1-year and 3-year survivals were similar to non-BCS cohort (84.2% vs. 94% and 71.3% vs. 91.9%, respectively, log rank test p = 0.09). CONCLUSION: LDLT is a good option for symptomatic BCS who have failed non-transplant interventions. The clinical and risk factor profile of BCS recipients is distinct from non-BCS recipients. By following an algorithmic management protocol, we show on propensity-score matched analysis that outcomes of LDLT for BCS are similar to non-BCS indications.

A study on drinking water quality in different income groups of Vizianagaram region of India.

Portable clean water consumption is the basic right of every individual. The major global concern is water pollution which can cause mortality. Change in physicochemical characteristics in drinking water is not only a pollution problem, but the presence of antibiotic-resistant microbes is also a significant issue. The study was carried out to assess the physicochemical and microbiological quality of the reservoir, municipality-supplied water, and bottled water. A total of 100 samples were collected from different income classes (higher, medium, and low) groups. The experiments were carried out based on the guidelines of APHA. In the present study, 40% of samples have been found to be contaminated with bacteria such as Escherichia coli, Vibrio cholerae, and Enterobacter aerogenes. Moreover, these bacteria also showed antibiotic resistance to certain drugs. The percentage of isolated bacterial strains was resistant to amoxicillin and ampicillin antibiotics. The statistical analysis of the Chi-square test states that there is a significant correlation between E. coli and other microbes (p ≤ 0.5). This study gives a piece of baseline information about the prevalence of antibiotic-resistant bacteria and focuses on the improvement of water from purification before reaching the consumer.

Maternal and Child Health Pipeline Training Programs: A Description of Training Across 6 Funded Programs.

PURPOSE: The HRSA-funded maternal and child health pipeline training programs (MCHPTPs) are a response to the critical need to diversify the MCH workforce, as a strategy to reduce health disparities in MCH populations. These MCHPTPs support students from undergraduate to graduate education and ultimately into the MCH workforce. DESCRIPTION: The models and components of training across the six MCHPTPs funded in 2016-2021 are summarized, to examine the design and delivery of undergraduate pipeline training and the insights gained across programs. ASSESSMENT: Strategies that emerged across training programs were organized into three themes: recruitment, support for student persistence (in education), and pipeline-to-workforce intentionality. Support for student persistence included financial support, mentoring, creating opportunity for students to develop a sense of belonging, and the use of research as a tool to promote learning and competitiveness for graduate education. Finally, the link to Maternal and Child Health Bureau (MCHB) long-term training and other MCHB opportunities for professional development contributed significant nuance to the pipeline-to-workforce objectives of these programs. CONCLUSIONS: The MCHPTPs not only increase the diversity of the MCH workforce, they also actively prepare the next generation of MCH leaders. The intentional connection of undergraduates to the infrastructure and continuum of MCH training, underscores the comprehensive impact of this funding.

A multicenter prospective observational study on the use of type and screen method versus conventional type and crossmatch policy for pre-transfusion testing in the Indian population.

Despite knowing the benefits of the type and screen (TS) method in pre-transfusion testing (PTT), most transfusion centers in developing countries continue to be reluctant to adopt a TS strategy over the conventional type and antihuman globulin (AHG) crossmatch (TX) policy in their routine laboratory practice because of the cost of obtaining antibody screening reagents. To generate strong evidence, this multicenter, observational study was conducted in which we collected data prospectively over a 1-year period from six major blood centers in India. The primary objective of this study was to identify the discordance between TS and TX results. A secondary objective was to identify the allo-antibody specificity in patients with positive antibody detection tests. All patients with orders for red blood cell transfusion who met patient selection criteria were subjected to parallel testing by column agglutination technology (CAT) for both the antibody detection test (screen) using a commercial three-cell panel and for the AHG crossmatch. A total of 21,842 patients were tested. In 148 patients with incompatible crossmatches, samples from six patients gave negative results with the antibody detection test, whereas the antibody detection test was positive in samples from 118 patients among the 21,694 crossmatch-compatible cases. The TS approach achieved a positive percent agreement of 95.95 and was found to be significantly effective in preventing the transfusion of serologically incompatible blood. The risk associated with abbreviating the AHG crossmatch was found to be 0.009 percent. Most of the identified clinically significant alloantibodies were directed to Rh antigens (D>E>c>C>e), followed by anti-K and anti-M. This study has generated sufficient robust data for the Indian population by including patients from all major geographical areas of the country and concluded a satisfactory agreement level as well as non-inferiority to the current PTT policy. Therefore, TS policy can be implemented in developing countries with no compromise on blood safety, provided sufficient technical and infrastructural support are available.

High-Frequency, Low-Dose Education to Improve Neonatal Outcomes in Low-Resource Settings: A Cluster Randomized Controlled Trial.

BACKGROUND: Annually 2.5 million infants die in the first 28 days of life, with a significant regional distribution disparity. An estimated 80% of those could be saved if neonatal resuscitation were correctly and promptly initiated. A barrier to achieving the target is the knowledge and skills of healthcare workers. PURPOSE: The objective of this cluster randomized trial was to assess the improvement and retention of resuscitation skills of nurses, midwives, and birth attendants in 2 birth centers serving 60 villages in rural India using high-frequency, low-dose training. RESULTS: There was a significant difference ( P < .05) between the groups in the rate of resuscitation, with 18% needing resuscitation in the control group and 6% in the intervention group. The posttest scores for knowledge retention at the final 8-month evaluation were significantly better in the intervention group than in the control group (intervention group mean rank 19.4 vs control group mean rank 10.3; P < .05). The success rate of resuscitation was not significantly different among the groups. IMPLICATIONS FOR PRACTICE: Improved knowledge retention at 8 months and the lower need for resuscitation in the intervention group support the efficacy of the high-frequency, low-dose education model of teaching in this setting. IMPLICATIONS FOR RESEARCH: Replication of these findings in other settings with a larger population cohort is needed to study the impact of such intervention on birth outcomes in low-resource settings.

First report of association of 'Candidatus Phytoplasma asteris' with Moringa oleifera leaf yellowing and stunting disease in India.

Moringa oleifera (family Moringaceae) also known as the 'drumstick tree' is a significant nutritious and medicinal plant that is commonly grown in India and contains a variety of vital phytochemicals. M. oleifera is used in several Indian herbal medicine formulations to treat a variety of illnesses (Kumar and Rao 2021). Typical phytoplasma symptoms of leaf yellowing and stunting were observed in M. oleifera trees up to 10% incidence at Acharya Narendra Dev University of Agriculture & Technology, Ayodhya, Uttar Pradesh, India in November 2021 and stunting with less fruit bearings symptoms with 8% incidence in October 2021 at Jonnalakothapalle village of Mudigubba mandal of Ananthapuramu district in Andhra Pradesh, India (Fig.1a, b). To investigate the possibility of a phytoplasma association with the symptoms, total DNA was isolated from the leaf samples collected from two diseased and two healthy plants from both the locations using CTAB method. The DNAs isolated were analysed by nested polymerase chain reaction (PCR) with universal phytoplasma primer pairs P1/P7 and R16F2n/R16R2 for the 16S rRNA gene (Deng and Hiruki 1991; Gundersen and Lee 1996) and secAfor1/sArev3 and SecAfor2/ SecArev3 for secA gene (Hodgetts et al. 2008). Amplicons of the expected size (~1.25kb from 16S rRNA gene and ~480bp from secA gene) were obtained from symptomatic plants only. The nested PCR products were cloned (pGEM-T Easy Vector, Promega), sequenced (ABA Biotech, India) and the sequences were deposited in GenBank with accession numbers OP358449, OP358450, OP358451, OP358452 for the 16SrRNA gene (~1.25 kb) and OP358443, OP358444, OP358445, OP358446 for the secA gene (~480 bp). BLASTn analysis revealed that the partial 16S rRNA gene sequences of M. oleifera phytoplasma isolate shared up to 99.9% sequence identity with the strain 'Candidatus Phytoplasma asteris' (Accession numbers MN909051, MN909047) and secA gene sequences shared up to 100% sequence identity with 'Ca. Phytoplasma asteris' (Accession numbers KJ434315, KJ462009) belonging to 16SrI group. The 16S rRNA and secA genes sequence-based phylogenetic analysis (Figure 1d,e) showed that the phytoplasma strain associated with M. oleifera leaf yellowing and stunting clustered within the 16SrI phytoplasma group closest to 16SrI-B ('Ca. P. asteris') subgroup strains. Furthermore, the virtual RFLP pattern derived from the query 16S rDNA F2nR2 fragment is identical (similarity coefficient 1.00) to the reference pattern of 16Sr group I, subgroup B (GenBank accession: AP006628). To the best of our knowledge, this is the first report of the 16SrI-B subgroup of the phytoplasma strains with M. oleifera in the world. 'Candidatus Phytoplasma asteris' (16SrI-B subgroup) strains have been reported from several other commercial crops and weed hosts in India and efficient leafhopper vectors have been identified (Rao 2021; Reddy 2021). This indicates that the 'Ca. P. asteris'-related strains (16SrI-B) are widespread and infecting several plant species in India. The increasing incidence of the 16SrI-B strain and its wide host range in India strongly suggests further research into the epidemiology involved in the dynamic spread of the disease in order to recommend a suitable management approach.

First Report of 'Candidatus Phytoplasma aurantifolia' Related Strain (16SrII-D) Associated with Stunting, Little Leaf and Phyllody Disease of Pearl Millet from South India.

Pearl millet [Pennisetum glaucum (L).R.Br.] also known as bajra, is one of the oldest millets and is cultivated in dry regions of arid and semi-arid tropics where no other cereal can be successfully grown. Pearl millet cultivation in India accounts for about two-thirds of millet production and is the fourth most cultivated food crop after rice, wheat and maize in India (Reddy et al. 2021a). In February 2021, the typical symptoms of stunting, phyllody and little leaf were observed after 25-30 days after sowing pearl millet seeds at Agricultural Research Station in Perumallapalle, Tirupati, India (Fig.1 A-C). The disease incidence was recorded up to 20% in the sampling regions. Total DNA was extracted from two symptomatic and two asymptomatic plant samples using CTAB DNA extraction method (Murray and Thompson, 1980). The extracted DNA was amplified in direct PCR and nested PCR assay using phytoplasma 16S rRNA universal primer pairs P1/P7 and R16F2n/R16R2 (Gundersen and Lee.1996) and secA gene with secAfor1/SecArev3 and SecAfor2/SecArev3 primer pairs (Hodgetts et al. 2008). 16SrRNA (1.25 kb) and secA (600 bp) gene amplicons were obtained from two symptomatic samples by nested PCR. No amplicons were produced with DNA from healthy leaf samples. Nested PCR amplified products (1.25 kb and 600 bp) from the symptomatic samples corresponding to the F2nR2 region of 16S rRNA and secA were directly sequenced at automated DNA sequencing facility (Eurofin Genomics India Pvt., Ltd Bangalore) and sequence data was deposited to NCBI GenBank with accession number ON005559 and ON067810. BLAST analysis revealed that pearl millet phytoplasma strain shared 100% sequence identity in 16Sr RNA and secA genes to 'Canditatus Phytoplasma aurantifolia' related strains (Acc. Nos. OM616883 and MT952965) from India. The subgroup was identified as 16SrII-D using the iPhyClassifier based on the virtual RFLP pattern derived from the query 16S rDNA F2nR2 fragment (Zhao et al. 2009). The virtual RFLP pattern is similar to the reference pattern of 16SrII-D (Y10096) with similarity coefficient 1.00. Phylogenetic analysis of 16S rRNA and secA gene sequences using MEGA version 7.0 revealed that the pearl millet phytoplasma strain clustered with 'Ca. P. aurantifolia' isolates of 16SrII-D subgroup. (Fig.1D-E) Earlier, one of 16SrI-B-phytoplasma strain (HM 134245) associated with green ear disease of pearl millet was reported in North India (Kumar et al. 2010). In this study, we reported the association of 16SrII-D subgroup phytoplasma with little leaves and witches'-broom disease of pearl millet in South India. Phytoplasmas belonging to the 16SrII-D subgroup have a wide range of hosts, including the agricultural and horticultural crops (Reddy et al., 2021b). Hence, this is the first report of 'Ca. P aurantifolia' infection in bajra in South India. The increase in the spread of 16SrII-D sub group phytoplasma diseases and the expansion of the host range strongly suggest further studies on the epidemiology of the dynamic dissemination of this disease in India.